During the growth and development of plants, unfavorable environmental conditions can be considered as environmental stresses, generally divided into biotic stress and abiotic stress. Biotic stress is caused by living organisms which can harm plants, such as viruses, fungi, bacteria, and harmful insects. Abiotic stress is caused by nonliving environmental factors that can have harmful effects on plants. This includes, for example, unfavorable conditions of water, temperature, salt, light, nutrition, wind, and the like.
The negative impact that the environmental stresses bring to the agricultural production is worldwide, and the impact caused by the abiotic stress, especially drought, is even worse (Boyer J S. 1982. Plant productivity and environment. Science 218:443-448). As Table 1 shows, abiotic stresses can reduce crop yields by 50% to 80%. Effort has been poured into the research of abiotic stress biology worldwide in order to discover key abiotic stress tolerance genes and unravel the molecular mechanisms underlying stress tolerance for crop improvement by gene engineering.
TABLE 1The average yield and the record high yield of eight different cropsTheThe average lossThe loss causedThe recordaverage(kg/ha)by abiotic stresshigh yieldyieldBioticAbiotic(% of record highCrop(kg/ha)(kg/ha)stressstressyield)maize19,3004,6001,95212,70065.8wheat14,5001,88072611,90082.1soybean7,3901,6106665,12069.3broomcorn20,1002,8301,05116,20080.6oat10,6001,7209247,96075.1barley11,4002,0507658,59075.4potato94,10028,30017,77550,90054.1beet121,00042,60017,10061,30050.7
Agriculture occupies a large sector in the economy of China, but abiotic stresses are major threats of Chinese agriculture, especially drought and salt stress being the two most important factors limiting sustainable agriculture. In China, 50% of arable farmland is affected by drought. Even in central and southern China with more rainfall, drought still occurs during the reproductive phase of rice, the predominant crop in the region and causes tremendous yield loss. In a severe drought, the entire crop yield may be lost. Insufficient rainfall in the vast area of northern and northwestern China makes the soil salinity problem even worse, which has become one of the major restraints of sustainable agriculture in the region.
The research of abiotic stress biology and cloning of stress tolerance gene are thus very important and urgent. Plant biologists worldwide work very hard to unravel the molecular mechanisms and isolate stress tolerance genes for crop improvement. Progress has been made, especially in the field of the molecular mechanism of salt stress by using the model plant Arabidopsis thaliana. Recently there were many important new discoveries (Zhu J K. 2002. Salt and drought stress signal transduction in plants. Annu. Rev. Plant Biol. 53:1247-273; Xiong L M, Schumaker K S, Zhu J K. 2002. Cell signaling during cold, drought, and salt stress. Plant Cell, S165-S183). For higher plants, sophisticated mechanisms have evolved to perceive the physical and chemical changes in their surrounding environment, and respond correspondingly by transducing the extracellular signals into the intracellular signals, and eventually relay the signals into nucleus and activate transcription factors that turn on corresponding gene expression, deploying their defense arsenals and adjusting their growth and development in order to adapt to the changed environment. Due to the complex nature of drought and salt stress effects, the molecular mechanisms underlying plant tolerance to these stresses are not fully understood although significant progress has been made (Ingram and Bartels 1996; Bray 1997; Bohnert 2000; Cushman and Bohnert 2000; Hasegawa et al. 2000; Bartels and Salamini 2001; Zhu 2002; Seki et al. 2003; Bray 2004; Amtmann et al. 2005; Zhang et al. 2005). The adaptive responses to these water deficit stresses must be coordinated at the molecular, cellular, and whole-plant levels. It is generally believed that roots first perceive a dehydration stress signal when the water deficit reaches a certain level. But how the physical signals of dehydration stress are perceived by the roots and converted into biochemical signals still remains unclear. Abscisic acid (ABA) is involved in coordinating whole plant responses since it is synthesized in the roots and translocated to the aerial portion of the plant, where it regulates stomatal behavior (Sauter et al. 2001). However, ABA receptors remained elusive until recently. One ABA receptor has finally been identified (Razem et al. 2006).The research on plant drought stress has been focused on the aspects of osmoregulation and the signal transduction of abscisic acid (ABA) and the loss-of-function mutants of Arabidopsis thaliana played an important role to this research (Zhu J K. 2002. Salt and drought stress signal transduction in plants. Annu. Rev. Plant Biol. 53:1247-273; Xiong L M, Schumaker K S, Zhu J K. 2002. Cell signaling during cold, drought, and salt stress. Plant Cell. S165-S183). Transpirational water loss through the stomata is an important determining factor of drought tolerance (Xiong et al. 2002 Plant Cell 14 Suppl: S165-183). Regulation of stomatal behavior has been an active area of research for drought stress, and much progress has been made (Luan 2002 Plant Cell Environ 25(2): 229-237). Root growth is another determining factor for drought tolerance (Malamy 2005 Plant Cell Environ 28(1): 67-77). Drought stress stimulates the growth of roots to extend to deeper soil to absorb water (Eapen et al. 2005 Trends Plant Sci 10(1): 44-50). Many studies have correlated root growth with drought tolerance (Zheng et al. 2000 Genome 43(1): 53-61). However, little is known about specific genes that are important for root growth under drought stress. Consequently, it has not been possible to engineer drought tolerance by enhancing root growth.
Genetic screening and analysis of loss-of-function mutants have helped us understand the plant stress tolerance mechanisms (Ishitani et al. 1997 Plant Cell 9(11): 1935-1949). However, for many genes, loss-of-function mutations do not lead to identifiable phenotypes due to functional redundancy. In addition, for some genes, loss-of-function mutations may be lethal. Gain-of-function mutants may overcome these shortcomings. Activation tagging is an effective method to generate gain-of-function mutants (Walden et al. 1994 Plant Mol Biol 26(5): 1521-1528; Weigel et al. 2000 Plant Physiol 122(4): 1003-1013).
The activation tagging method has been used successfully in identifying a number of gain-of-function mutants in plant development or hormonal responses (Kakimoto 1996; Kardailsky et al. 1999 Science 286(5446): 1962-1965; Borevitz et al. 2000 Plant Cell 12(12): 2383-2394; Ito and Meyerowitz 2000 Plant Cell 12(9): 1541-1550; Lee et al. 2000 Genes Dev 14(18): 2366-2376; van der Graaff et al. 2000 Development 127(22): 4971-4980; Huang et al. 2001 Plant Physiol 125(2): 573-584; Zhao et al. 2001 Science 291(5502): 306-309; Razem et al. 2006 Nature 439(7074): 290-294). However, despite the power of the activation tagging approach, it has not been adequately explored for drought tolerance studies. Only a few activation tagged gain-of-function mutants with enhanced abiotic stress tolerant phenotype have been reported (Furini et al. 1997 Embo J 16(12): 3599-3608; Ahad et al. 2003 Transgenic Res 12(5): 615-629; Grant et al. 2003 Mol Plant Microbe Interact 16(8): 669-680; Aharoni et al. 2004 Plant Cell 16(9): 2463-2480; Chini et al. 2004. Plant J 38(5): 810-822). In contrast to the loss-of-function mutant, the gain-of-function mutants have not been adequately explored although the mutants can provide valuable materials for stress tolerance gene discovery and the cloned gene can be directly used for crop improvement.